Bioactives are documented to exhibit diverse pharmacological activities, however, their low and inconsistent bioavailability primarily pose serious impediment against their potential therapeutic usage. Efforts, therefore, have been undertaken to systematically develop nanostructured lipidic carriers of chrysin, a vital flavonoid, employing Capmul PG-12 (i.e., liquid lipid), glyceryl monostearate (i.e., solid lipid), stearylamine, Phospholipid S-100 (i.e., cosurfactant) and Poloxamer 188 (i.e., surfactant). NLCs were formulated using hot-melt dispersion-high pressure homogenization method and optimized using Face-Centred Cubic Design. Afterwards, stearylamine was conjugated with biotin as ligand through EDC-NHS coupling reaction and biotin-staerylamine complex formation was ratified using H-NMR and FTIR. It was further used instead of SA for the preparation of biotin-conjugated-optimized NLCs (Bio-NLCs). Mean particle size of consequent Bio-NLCs was found to be 246.4 nm and zeta potential as 11.4 mV. In vitro release studies indicated sustained drug release characteristics from NLCs over 48 h. Cell line studies conducted on coumarin6-loaded Bio-NLCs in demonstrated remarkably superior cellular uptake over naive NLCs and pure dye. Marked improvement in absorption parameters was observed during in vivo pharmacokinetics for Bio-NLCs and NLCs vis-à-vis pure chrysin suspension. However, the improvement for naive NLCs was relatively lower than that of Bio-NLCs. Almost 5.20-folds augmentation in C (p < 0.005), 8.94-folds in AUC (p < 0.001), 7.46-folds in AUC (p < 0.001) and 7.25-folds in Ka (p < 0.01), signify improved degree of drug absorption and retention of Bio-NLCs. Stability studies indicated the robustness of Bio-NLCs, when stored under refrigerated storage conditions for 3 months. By and large, the current work demonstrates high potential of Bio-NLCs for distinctly improved biopharmaceutical performance of chrysin.